Project Abstract Polymeric repeats, typically consisting of only one or two different amino acids, are found within many fungal proteins, particularly those with regulatory functions. These ?polyX? regions can be highly polymorphic, a consequence of slippage between repeat units during DNA replication. The general hypothesis to be tested here is that polyX polymorphisms fill the adaptation gap between conventional mutation and epigenetics. The specific focus of this R21 is the opportunistic yeast Candida glabrata, well known for its ability to develop resistance to antifungal azoles and, more recently, echinocandins. While single base mutations within C. glabrata Pdr1 or Fks proteins mediate high level resistance to these agents, there is emerging evidence that these mutations are accompanied by other genetic changes that confer reduced antifungal susceptibility or tolerance, or compensate for the decreased fitness associated with Pdr1/Fks mutation. In Aim 1, the polymorphism of 36 polyX proteins relevant to azole or echinocandin susceptibility will be assessed in 7 azole resistant and 6 echinocandin-resistant clinical isolates relative to their epidemiologically or phylogenetically matched susceptible isolates. Preliminary studies have identified one such polymorphism in the polyQ region of transcriptional repressor Tup1A that correlates with 16 to 32-fold reduced echinocandin killing. Aim 2 will directly test these correlations by using gene replacement to construct strains that differ only in the polyX content of an antifungal-relevant protein, followed by susceptibility and fitness assays. If the general hypothesis is supported, future studies would be warranted to evaluate the role of polyX polymorphism in other aspects of fungal infection, including tissue-specific adhesion, biofilm formation, and immune evasion.